褪黑素通过调节胶质瘤细胞中microRNA-155的表达发挥抑癌作用的机制研究
发布时间：2018-07-13 17:34
【摘要】：【目的】 检测经褪黑素处理后的胶质瘤细胞株中microRNA-155(miR-155)的表达情况，探讨褪黑素对胶质瘤细胞增殖、转移和侵袭能力的影响，并探究其抑癌的相关机制。 【方法】 第一部分：1、选用人脑胶质瘤细胞株U87、U373、U251共三株，分别用不同浓度（100μM、1μM、1nM）的褪黑素处理。运用qRT-PCR技术检测三株胶质瘤细胞中miR-155的表达量相对于对照组的变化。明确褪黑素和miR-155表达量的相关性，并确定引起miR-155表达量变化最大的褪黑素浓度。2、选定一株胶质瘤细胞株，并选用引起miR-155表达量变化最大的浓度范围的褪黑素处理细胞。通过CCK-8方法观察胶质瘤细胞的增殖情况。3、通过流式细胞仪技术检测胶质瘤细胞增殖、细胞周期及凋亡的变化。4、利用Transwell试验检测褪黑素对肿瘤细胞迁移、侵袭的影响。 第二部分：1、选定一株胶质瘤细胞，并选取引起miR-155表达量变化最大的浓度范围的褪黑素处理细胞。分别应用Western blot和qRT-PCR技术检测细胞中MYB基因的mRNA及蛋白表达相对于对照组的变化，明确褪黑素与MYB基因的相关性。2、选同一株胶质瘤细胞，将MYB基因干扰后用qRT-PCR技术检测细胞中miR-155的表达量相对于对照组的变化，明确MYB和miR-155的相关性。 【结果】 第一部分：1、三株胶质瘤细胞U87、U373、U251经过不同浓度（100μM、1μM、1nM）褪黑素处理后，miR-155的表达相对于正常对照组均明显下降(P0.05)，其中用1μM浓度褪黑素处理的细胞株中miR-155表达下降最明显(P0.01)。2、经过1μM浓度褪黑素处理的U87胶质瘤细胞增殖较对照组明显下降。3、经过1μM浓度褪黑素处理的U87胶质瘤细胞凋亡较对照组增高，而细胞周期没有变化。4、经过1μM浓度的褪黑素处理的U87胶质瘤细胞迁移和侵袭能力较对照组明显下降。 第二部分：1、经过1μM浓度褪黑素处理的U87胶质瘤细胞中MYB基因的mRNA和蛋白表达量均明显低于对照组。2、在U87胶质瘤细胞中干扰MYB基因后，miR-155的表达量下降。 【结论】 本研究结果表明褪黑素通过下调MYB基因从而抑制miR-155表达，1μM浓度褪黑素可以增加肿瘤细胞的凋亡使胶质瘤细胞增殖降低，并且能抑制胶质瘤细胞的迁移和侵袭能力，，进而发挥抑癌作用。本实验阐明了褪黑素在脑胶质瘤中抑癌作用的microRNA表观调控机制，为揭示褪黑素的抗脑胶质瘤作用机制奠定了理论基础，为发现脑胶质瘤的临床治疗靶点提供新思路。
[Abstract]:[objective] to detect the expression of microRNA-155 (miR-155) in glioma cells treated with melatonin, and to explore the effect of melatonin on the proliferation, metastasis and invasion of glioma cells. In the first part, three human glioma cell lines, U87U373U251, were treated with melatonin at different concentrations (100 渭 M, 1 渭 M, 1nM), respectively. The expression of miR-155 in three glioma cells was detected by qRT-PCR. To determine the correlation between the expression of miR-155 and melatonin, and to determine the concentration of melatonin. 2. To select a glioma cell line, and to select the melatonin treated cells in the concentration range that caused the most change of miR-155 expression. The proliferation of glioma cells was observed by CCK-8 method, the proliferation, cell cycle and apoptosis of glioma cells were detected by flow cytometry, and the effects of melatonin on the migration and invasion of glioma cells were detected by Transwell test. The second part: 1, a glioma cell line was selected, and melatonin treated cells were selected in the concentration range that caused the greatest change of miR-155 expression. The mRNA and protein expression of MYB gene were detected by Western blot and qRT-PCR. The correlation between melatonin and MYB gene was determined, and the same glioma cell line was selected. The expression of miR-155 was detected by qRT-PCR after MYB gene interference compared with the control group. The correlation between MYB and miR-155 was clarified. [results] in the first part, the expression of melatonin in U87U373U251 glioma cells was significantly lower than that in the control group (P0.05) after being treated with melatonin at different concentrations (100 渭 MU 1 渭 M) (P0.05). The expression of miR-155 in U87 glioma cells treated with 1 渭 M melatonin was significantly decreased (P0.01). The proliferation of U87 glioma cells treated with 1 渭 M melatonin was significantly lower than that of the control group. The apoptosis of U87 glioma cells treated with 1 渭 M melatonin was higher than that of the control. The cell cycle of U87 glioma cells treated with melatonin (1 渭 M) was not changed. The migration and invasion ability of U87 glioma cells treated with 1 渭 M melatonin was significantly lower than that of the control group. In the second part, the mRNA and protein expression of MYB gene in U87 glioma cells treated with melatonin at 1 渭 M concentration was significantly lower than that in control group, and the expression of miR-155 in U87 glioma cells was decreased after interfering with MYB gene in U87 glioma cells. [conclusion] this study suggests that melatonin can increase the apoptosis of tumor cells and decrease the proliferation of glioma cells by down-regulating the expression of miR-155 and inhibiting the expression of miR-155. And can inhibit the migration and invasion of glioma cells, and then play a role in cancer suppression. In this study, the mechanism of microRNA apparent regulation of melatonin in gliomas was elucidated, which laid a theoretical foundation for revealing the mechanism of melatonin's anti-glioma, and provided a new idea for the clinical treatment of glioma.
【学位授予单位】：苏州大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R739.4
本文编号：2120191
[Abstract]:[objective] to detect the expression of microRNA-155 (miR-155) in glioma cells treated with melatonin, and to explore the effect of melatonin on the proliferation, metastasis and invasion of glioma cells. In the first part, three human glioma cell lines, U87U373U251, were treated with melatonin at different concentrations (100 渭 M, 1 渭 M, 1nM), respectively. The expression of miR-155 in three glioma cells was detected by qRT-PCR. To determine the correlation between the expression of miR-155 and melatonin, and to determine the concentration of melatonin. 2. To select a glioma cell line, and to select the melatonin treated cells in the concentration range that caused the most change of miR-155 expression. The proliferation of glioma cells was observed by CCK-8 method, the proliferation, cell cycle and apoptosis of glioma cells were detected by flow cytometry, and the effects of melatonin on the migration and invasion of glioma cells were detected by Transwell test. The second part: 1, a glioma cell line was selected, and melatonin treated cells were selected in the concentration range that caused the greatest change of miR-155 expression. The mRNA and protein expression of MYB gene were detected by Western blot and qRT-PCR. The correlation between melatonin and MYB gene was determined, and the same glioma cell line was selected. The expression of miR-155 was detected by qRT-PCR after MYB gene interference compared with the control group. The correlation between MYB and miR-155 was clarified. [results] in the first part, the expression of melatonin in U87U373U251 glioma cells was significantly lower than that in the control group (P0.05) after being treated with melatonin at different concentrations (100 渭 MU 1 渭 M) (P0.05). The expression of miR-155 in U87 glioma cells treated with 1 渭 M melatonin was significantly decreased (P0.01). The proliferation of U87 glioma cells treated with 1 渭 M melatonin was significantly lower than that of the control group. The apoptosis of U87 glioma cells treated with 1 渭 M melatonin was higher than that of the control. The cell cycle of U87 glioma cells treated with melatonin (1 渭 M) was not changed. The migration and invasion ability of U87 glioma cells treated with 1 渭 M melatonin was significantly lower than that of the control group. In the second part, the mRNA and protein expression of MYB gene in U87 glioma cells treated with melatonin at 1 渭 M concentration was significantly lower than that in control group, and the expression of miR-155 in U87 glioma cells was decreased after interfering with MYB gene in U87 glioma cells. [conclusion] this study suggests that melatonin can increase the apoptosis of tumor cells and decrease the proliferation of glioma cells by down-regulating the expression of miR-155 and inhibiting the expression of miR-155. And can inhibit the migration and invasion of glioma cells, and then play a role in cancer suppression. In this study, the mechanism of microRNA apparent regulation of melatonin in gliomas was elucidated, which laid a theoretical foundation for revealing the mechanism of melatonin's anti-glioma, and provided a new idea for the clinical treatment of glioma.
【学位授予单位】：苏州大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R739.4
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